Chronic pain following acute peripheral nerve injury is a major public health issue. Our primary goal in this proposal is to understand the cellular processes that underlie plasticity in cutaneous and muscle sensory neurons following injury and regeneration. Specifically, we will look at injury-induced plasticity of primary sensory neurons reinnervating either skin or striated muscle. In the first aim, we use ex vivo skin or muscle/nerve/DRG/spinal cord preparations and our novel method for recovering individual functionally characterized sensory neurons to determine the comprehensive phenotypes of cutaneous C-fibers and Group IV muscle afferents in naive mice. This includes characterization of the gene expression profiles of the individual functionally characterized fibers. In our preliminary studies we have indentified several unique functional classes of these fibers. In the second aim, we will first examine the effects of nerve injury on the functional properties and correlated changes in gene expression in individual characterized fibers reinnervating skin or muscle. In our preliminary studies we have confirmed and expanded upon earlier studies showing that these fibers are sensitized following regeneration. In addition, our results suggest that some fibers are changing phenotype. Next we will examine changes in DRG expression of specific trk, GFRalpha, ASIC, TRP and purinergic receptors/channels contained in these fibers to indentify possible molecular mechanisms responsible for the changes in function observed following nerve injury and regeneration. In the third aim, we will use our novel in vivo siRNA knockdown technique to test the hypothesis that the injury-induced increase in the expression of particular receptors/channels (e.g. ASIC, TRP and purinergic) mediates the observed functional plasticity in specific populations of cutaneous and muscle afferents. For example, we expect to find that knockdown of the increase in TRPM3 expression levels following cutaneous nerve injury and regeneration will block the observed sensitization of cutaneous C-polymodal fibers to heat. Similarly we expect that knockdown in ASIC3 expression following muscle nerve injury and regeneration will block the sensitization of muscle afferents to mechanical stimulation. The determination of the specific receptors/channels responsible for sensitization of these fibers will provide new insights to these processes and more importantly could provide potential targets for the development of pharmaceutical therapies. These new therapies could provide for improved treatments for the alleviation of the adverse symptoms of chronic neuropathic pain.